A recombinant vaccinia virus which expresses glycoprotein D of HSV-1 has been tested for its efficacy as a vaccine to prevent primary and latent infections with HSV-1 and HSV-2. Mice vaccinated with the recombinant virus were challenged with a lethal dose of HSV-1 or HSV-2. Over 90% of the mice were protected in both cases. To determine whether vaccination would prevent the development of latent HSV infection, mice were vaccinated intradermally (tail route) with vaccinia-HSV-1 gD recombinant and four weeks later challenged with HSV-1 (KOS) by the lip route. Approximately two-thirds of the ganglia from vaccinated mice wee protected from the development of a latent infection, in contrast to less than 5% protection in the control mice. This is the first demonstration of the effectiveness of a genetically engineered vaccine to prevent the establishment of a latent HSV infection. Using in situ hybridization techniques, the presence of HSV gene transcript could be demonstrated in trigeminal ganglia of latently infected mice. To identify the gene(s) expressed, polyadenylated mRNA from trigeminal ganglia of latently infected mice was used as a template to synthesize labeled cDNA molecules which could be used as probes for Southern blots of cloned HSV DNA restriction fragments. mRNA samples were obtained from mice of ages ranging up to 24-month old animals and mRNA from trigeminal ganglia of uninfected mice was used as a control. Probe sequences capable of hybridizing with some viral fragments but not with others wre found in the samples from animals of all ages but not in the controls. The viral fragments detected were always the same, regardless of the age of the mice used, and finer mapping of the restriction fragments involved indicates that transcription of the viral genome during latency is restricted to two, possibly three of the HV immediate early genes.